Purification and biochemical properties of SDS-stable low molecular weight alkaline serine protease from Citrullus colocynthis

Abstract

A low molecular weight serine protease from seeds of Citrullus colocynthis was purified to electrophoretic homogeneity with high level of catalytic efficiency (22,945 M⁻¹ S⁻¹). The enzyme was a monomer with molecular mass of 25 kDa estimated by SDS–PAGE. The enzyme was highly active over a pH range of 6.5–9.0 and temperature range of 20–80 °C, with maximum activity at pH 7.5 and at 50 °C. The K_m and K_cat were 73 μg/mL and 67/s, respectively. The enzyme was strongly inhibited by PMSF, moderately by soybean trypsin inhibitor, indicating that the enzyme was a serine protease. The enzyme retained 86 and 73% of its activity in the presence of urea and DTT, respectively, and its activity was slightly enhanced in the presence of anionic detergent (SDS). Thus, the enzyme is a novel SDS-stable protease with high catalytic efficiency over wide ranges of pH and temperature which is commercially promising for various industrial applications.

Graphical abstract

Keywords:

• Citrullus colocynthis
• serine protease
• purification
• biochemical characterisation
• thermostability
• detergent stability

Acknowledgement

We are very grateful to Prof. Muhammad Iqbal Chaudry, director of HEJ-Research Institute of Chemistry, University of Karachi for allowing us to use their research facilities.

Disclosure statement

No potential conflict of interest was reported by the authors.